Project Summary: Clear neuroadaptations occur as a result of alcohol use disorders, underlying the debilitating health risks and social problems associated with the disease. While changes in GABAergic, glutamatergic and dopaminergic neurotransmission are strongly implicated, the precise neurological mechanisms that cause enhanced susceptibility of alcohol intake and preference are not yet well understood. As a result, current treatments available to combat alcoholism are sparse and nonspecific? besides psychotherapy, detoxification and nausea-inducing, emetic drugs, there are no pharmacological targets for treating the disease. Recent studies have uncovered non-canonical GABA synthesis mediated by aldehyde dehydrogenase 1a1 (ALDH1a1) and co-release from dopaminergic midbrain neurons, and we have gathered strong evidence suggesting that this alternative GABA synthesis pathway is diminished with blood alcohol levels that are associated with binge drinking. Methodological advances now provide the opportunity to directly examine and dissect the role of this GABA pathway in acute and chronic alcohol exposure. In this proposal, we hypothesize that the balance between co-released dopamine and GABA by midbrain dopamine neurons is critical for postsynaptic striatal spiny projection neurons (SPNs). In addition, alcohol and/or its metabolic product inhibit dopaminergic GABA by impairing GABA non-canonical synthesis, ultimately perturbing the co-released GABA by dopamine neurons in the dorsal striatum, and lead to enhancement in susceptibility of alcohol intake and preference. We will employ the use of genetically modified mice and virus injections to directly examine dopamine neurons and the specific pathways involved in GABA synthesis, modulation and release. By optogenetic activation of dopamine terminals in the striatum and whole-cell patch clamp on identified principle striatal cells, our experiments will provide a high-resolution assessment of the function of dopaminergic GABAergic in normal and enhanced alcohol intake and preference conditions. Combined with imaging, electrophysiology, optogenetics, biochemistry and behavioral analysis, this proposal provides a unique, multi- faceted approach to study dopamine and GABA co-release in the basal ganglia, and the mechanisms by which chronic binge drinking alters this form of neurotransmitter release. Based on our compelling preliminary findings, we aim to 1) examine whether cell-type specific deletion of ALDH1a1 in dopamine neurons leads to enhanced alcohol preference; 2) pinpoint the mechanism underlying the inhibition of co-released GABA by alcohol; and 3) investigate alcohol modulation of functional interplay between GABA and dopamine in the striatum. Our novel initial findings and the studies proposed in this grant will close the gaps in our knowledge about GABA synthesis in dopaminergic neurons and create a new window into our understanding of synaptic mechanisms underlying enhanced alcohol intake and preference, providing better tools and earlier targets for the treatment or prevention of alcohol use disorders. !